Haemophilus influenzae is a pathogen that mediates a wide range of human disease including otitis media, sinusitis, and pneumonia often in association with an increased density of nasopharyngeal colonization. As is the case with other bacterial respiratory pathogens, an epidemiologically relevant risk factor for this process is prior injury of the respiratory mucosa associated with respiratory viral infections and environmental irritants (e.g. smoking). Colonization of the upper respiratory tract is an essential event in the pathogenesis of disease and is facilitated by bacterial adhesins that recognize specific receptors motifs on eukaryotic cells. The hemgglutinating pilus is perhaps the most well- studied H. influenzae adhesin and is expressed by both non-typable and serotypable strains. Our preliminary evidence suggests that this structure mediates adherence to damaged and extruding non-ciliated respiratory epithelium. A two-fold hypothesis to be pursued in this proposal is that this microbial virulence factor functions to compromised areas of the respiratory mucosa for subsequent pathogenic events and then further enhances the rupture of mucosal defenses by inducing respiratory epithelial cell apoptosis. Using the H. influenzae pilus as a model system, we will dissect the multiple roles in specific pilus receptor structure(s) on human respiratory epithelium by using piliated/non-piliated isogenic strains and microsphere-linked pilus adhesin probes. We will then use these tools to characterize the dynamics of pilus receptor distribution on a re-organizing mucosal surface to determine whether this adhesion recognizes focal areas associated with its receptor can result in the manipulation of eukaryotic cell signal transduction mechanisms, increases the apoptotic index of epithelial cells, and disrupts the barrier of tight junctions between cells. The dynamics of respiratory colonization to surmount mucociliary clearance and cause disease. By focusing on this microbial virulence factor, we will be able to develop broadly applicable experimental systems that can be used to dissect the molecular details of host-pathogen interactions in respiratory disease.